Identification of CCR8 as the specific receptor for the human beta-chemokine I-309: cloning and molecular characterization of murine CCR8 as the receptor for TCA-3.
Chemokine receptor-like 1 (CKR-L1) was described recently as a putative seven-transmembrane human receptor with many of the structural features of chemokine receptors. To identify the ligand of CKR-L1, we have studied chemokine-induced calcium mobilization in 293 cells transfected with CKR-L1. Of 20 different chemokines tested, only I-309 was able to elicit a significant calcium mobilization. In addition, I-309 induced the transfectants to migrate in vitro. As expected for chemokine receptor-mediated effects, pertussis toxin, but not cholera toxin, inhibited both the calcium flux and migration of the CKR-L1 transfectants in response to I-309. All of these data support the conclusion that I-309 is a functional ligand for CKR-L1. According to the current chemokine receptor nomenclature, we have designated this gene as CCR8. The murine CCR8 (mCCR8) gene was cloned, and its predicted amino acid sequence showed a 71% identity with that of human CCR8. As human CCR8, mCCR8 is expressed in thymus. Both I-309 and its murine homologue TCA-3 were able to induce calcium mobilization in transiently transfected 293-EBNA cells expressing mCCR8. The affinity of the binding of 125I-labeled TCA-3 to mCCR8 was high (Kd approximately 2 nM); the binding was prevented completely by an excess of cold TCA-3, and only partially competed (40%) by I-309. The identification of I-309 and TCA-3 as the functional ligands for CCR8 receptors will help to unravel the role of these proteins in physiologic and pathologic situations.
Goya
I
I
Departamento de Inmunología y Oncología, Centro Nacional de Biotecnología, Madrid, Spain.
Gutiérrez
J
J
Varona
R
R
Kremer
L
L
Zaballos
A
A
Márquez
G
G
D000595:Amino Acid Sequence; D000818:Animals; D002118:Calcium / Q000378:metabolism; D002460:Cell Line; D054402:Chemokine CCL1; D019742:Chemokines, CC / Q000378:metabolism* / Q000502:physiology; D002633:Chemotaxis / Q000187:drug effects; D003001:Cloning, Molecular; D016207:Cytokines; D006801:Humans; D007457:Iodine Radioisotopes; D007668:Kidney / Q000166:cytology; D051379:Mice; D008969:Molecular Sequence Data; D054401:Receptors, CCR8; D019707:Receptors, Chemokine / Q000096:biosynthesis / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D018121:Receptors, Cytokine / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D014162:Transfection
C065971:CCL1 protein, human; C103330:CCR8 protein, human; C502258:Ccl1 protein, mouse; C489319:Ccr8 protein, mouse; D054402:Chemokine CCL1; D019742:Chemokines, CC; D016207:Cytokines; D007457:Iodine Radioisotopes; D054401:Receptors, CCR8; D019707:Receptors, Chemokine; D018121:Receptors, Cytokine; D002118:Calcium